Download Restriction Enzyme

From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
Factor
A
cDNA
for
investigate
has
of
absent
in
in
in
the
paternal
normal
XII
factor
of molecular
weight
inactive
proenzyme;
protease
in the
is
production,
when
and
tomatic
hereditary
a low
clotting
report
anomaly
5’ portion
11
as
trait
a
is
to an apparent
the
the
cause
1987
by Grune
DNA
sulfate
Coagulation
on
in
first
The
study
and
of
the
markedly
and
brothers
prolonged
normal
bleeding
Specific
deficiency
lack of factor
XII
RFLP
detection.
distribu-
locus, useful
for
and for genetic
antibody
specific
indicates
that
length
polymorphisms
the inheritance
tracing
linkage
studies.
In this paper
carrying
(RFLPs)
at the
of Hageman
a FXII
Hageman
TaqI
trait
and
lineage,
is reported.
This
FXII gene and correlated
the
family
RFLP,
in
RFLP
with
XII
activity
was
using
antigen
was
clotting
an
has
Southern
unrelated
of
the
of
partial
deficient
by
quantitative
technique.’
used
blot
hybridization.
family
Blood
under
FXII
monospecific
AG,
different
time
substrate.’
immunoelectrophoresis
A
(Behringwerke
from
thromboplastin
plasma
samples
Italian
study
from
were
obtained
35
and
normal
from
by
standard
formed
with
as previously
restriction
England
FXII
library
Blood,
and
the
Southern
described.’0
expression
Vol 69, No
5 (May),
technique
Six to 10 g of DNA
endonucleases
(Boehringer
Biolabs, Boston).
cDNA
probes were
in the
blotting
Mannheim,
were
per-
were digested
FRG;
New
obtained
i987:
pEXI.8
pp
from
a human
All
experiments
i42i-i424
liver
cDNA
involving
IV-2
hemostatic
showed
and
TaqI
Roma;
thrombin
the
ofthe
with
probes
The
partial
digests
two
DNA
di Studi
Centro
several
delle
Address
span
sul
The
lo Studio
inserts
the
2A)
two
as
An
brothers
di Cooley,
Umana,
e Ia
121
four subjects
examined.
two
Morbo
leader
aminoacid
2 (Fig
in the
sul
and
Universi-
Universit#{224} di
Terapia
1 1, /986;
accepted
Progetto
Finalizzato
Malattie
dell’Emofilia,
Ricerche
1 1, 1986.
Genetica
contratto
e
Consiglio
n. 8400877.
requests
Morbo
December
Ingegneria
Ereditarie
di
to
F.
Bernardi,
Cooley.
Via
BS,
Luigi
Centro
Borsari,
di
46.
Studi
44100
Italy.
publication
costs
ofthis
article
payment.
This
article
must
charge
“advertisement”
©
found
cDNA
to an
ofthe
from
from
di Biopatologia
delle
reprint
Biochimici
restriction
Italy.
September
by the
Molecolari
1) and
for FXII.
to probe
Biochimici
per
Universit#{224} di Pisa,
with
in Fig
sequence
cDNAs
was
alterations
from a human
liver
by immunoblotting
hybridized
band
times.
severe
and a
brothers
pedigree
digested
Dipartimento
and
time
showed
a
C < I U/dL)
from
isolated
vector)
a
thromboplastin
to cDNA
for FXII.
the
Centro
indicate
vector
and
have been
expression
t#{224}
di Ferrara;
Ferrara,
isolation
(New
trait
factors
(XII:
DNA
was
2.1-kb
Nazionale
24
procedures.
DNA
method
bands
of 3.3 and 1.1 kb were found
in the
well as in the 35 unrelated
normal
individuals
Basi
FRG).
regions
10%
in accor-
The
prothrombin
hybridized
Submitted
Supported
heterologous
Marburg,
were
assays.
to aminoacid
79 (probe
1) and
(probe
2) of plasma
FXII.8
From
FXII
determinations.
safety
Hageman
partial
gene,
parents
In the
paternal
within
activities
METHODS
antigen
by the activated
been
the
their
peptide
to 396
brothers
of their
has been mapped
the FXII plasma
and
appropriate
Laurell’s
subjects
members
1 1 members
AND
activity
estimated
to
antibody
in two
from
additional
determined
method
according
detected
fragment
members.
MATERIALS
Factor
FXII
trait
enables
filters
antigen
(XII: Ag < 10 U/dL).
In order to investigate
gene
probes
(pEXI
restriction
haploid
and
membrane
with
and
The
library
FXII7’8
of
Data
per
formamide
of the
assays
for coagulation
of FXII clotting
activity
the
for
deficiency.
gene
to the accepted
(50%
activated
and
and
XII
according
immunologic
two
enzymes
cDNA
the
site
a
reported.6
of
within
RESULTS
to
also
lesions
has
polymorphic
XII
transfer
asymp-
(IV-l
gene
factor
washing
screen
trait
cloning
localized
in the
factor
family
constructed.
Inc.
at 42#{176}C)and
Hageman
of
the
one
been
been
conditions
FXII
recent
has
were performed
Hybridization
ofthe
The
of
of
in this
has
mutation
& Stratton,
of RFLPs
investigation
trait
map
The
presence
tion, Veltkamp
et a15 inferred
that at least two allebomorphic
genes control
the synthesis
of the factor.
A different
form of
Hageman
trait
with autosomal
dominant
inheritance
has
been
Hageman
site
gene.
dance
with the gene
England
Nuclear).
a complete
a partial
bimodal
probably
dextran
character
trait)
causes
in heterozygotes,
of
restriction
polymorphic
of the
recombinant
kinin
factor
recessive
A
TaqI
guidelines.
serine
assays.
Since
Ratnoff’s4
of cases
have been observed.
Due
a)
based
of the
cause
gene
due
diagnosis
antigen
the
genome.
as an
is an
is
the
suggest
the
cascade,
coagulation
its
and
(Hageman
activity3
and
of activity.
acts
clotting
as an autosomal
homozygotes
of FXII
deficiency
FXII3;
hundreds
is transmitted
that
in
deficiency
and
identifica-
in plasma
it
blood
blood
activity
immunologic
( 1 954),
with
in
XII
as
A. Fantoni,
and F. Conconi
excluded.
frag-
with
the
Factor
Hageman
of
plasma
and
activated,
in vitro
deficiency
finding
correlated
enables
circulates
of the
of
and
deletion
Detected
M. Tripodi,
F. Panicucci,
been
factor
XII or Hageman
facwith a single
polypeptide
fibrinolysis.2
anomaly
to
polymorphism.
deficiency.
80,000,’
initiation
brothers
polymorphic
This
and
XII
used
Trait
L. del Senno,
L. Rossi,
restriction
propositi
lineage.
activity
been
two
two
P. Patracchini,
L. Felloni,
and
Taql
population.
of factor
has
lesions
A
LOOD
COAGULATION
tor (FXII),
a glycoprotein
chain
XII
gene
family.
found
of heterozygous
B
factor
of
their
the
the
reduction
tion
and
been
members
F. Vannini,
polymorphisms
trait
ment
G. Marchetti,
S. Bartolai,
presence
length
Hageman
Gene
Alteration
in Hageman
by TaqI
Restriction
Enzyme
By F. Bernardi,
coagulation
the
fragment
XII
/987
in
accordance
with
were
defrayed
therefore
18
U.S.C
be
in part
hereby
§1734
by page
marked
solely
to
this fact.
by Grune
& Stratton,
Inc.
0006-4971/87/6905-0023$3.00/0
142 i
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
BERNARDI
i 422
ET AL
III
IV
._.w
,R
Fig 1 .
Pedigree
of the Hageman
activity
(FXII:C),
when determined,
their
father.
detected
The
polymorphic
in hybridizations
Family
Figure
results
of
been
1 shows
pedigree
the
RFLP
lineage;
intermediate
(Fig
5’ cDNA
of the
studied
analysis
TaqI
RFLP
is present
subjects
in seven males
the
inheritance
activity
have
trait subjects.
the
band
with
The
studies.
the FXII
Hageman
trait family. . presence
of Taql polymorphism;
is reported
below each member
of the family.
in the
2B)
was
probe
I.
TaqI
family
of the family
and
of the FXII
activity
levels
normal
control
values
are
in the polymorphic
RFLP
reported
have
localize
in the
been
FXII
Fig
3;
originates
be present
the
The
BamHI
nonpolywith
IS
activities
are
gene
localization.
to determine
the
Taql
Additional
the
origin
and
produced
analysis.
RFLP.
B
H
an intronic
DNA
lit
S...
..#{149}
*
that
band;
diploid
the
SB).
3.3
cDNA
was
that the
detectable
polymorphic
pattern
probes,
present
(data
polymorphic
thus two
genome.
band
FXII genes
Hybridizations
should
of
(Fig 4)
polymor-
polymorphism
by Southern
is therefore
to localize
double
the
digestions
TaqI
with
polymorphic
several
origiblot
explained
In Fig 5A NcoI/TaqI
patterns
are reported.
(BclI/TaqI)
TaqI
polymorphic
fragments
Exonic
by
with
in the
and BclI/TaqI
Two polymorphic
additional
detected
RsaI)
by the
FXII
enzymes
and of 1.2-kb
(NcoI/TaqI)
subject.
Analysis
of the
obtained
sequences
site
restriction
(TaqI/BglII,
HindIII,
PstI, PvuIl,
TaqI polymorphic
site in the 5’ portion
Kb
-
suggests
both
band
sequence.
performed.
digestion
in a
-1
using
2. 1 -kb
thus excluding
a gene
deletion
The
0.47-kb
A
band,
the
an additional
TaqI site cleaving
the 3.3-kb
DNA
fragment
in
the 2.1-kb
band and in a 1.2-kb
band not hybridizing
to the
cDNA
probes
used. This I .2-kb DNA
fragment
is probably
gene,
to
3.3-kb
lineage.
FXII
subjects.
and BglII
DNA
digests
with both probes
large FXII bands common
to normal
and
phic subjects,
nates
from
in
carried
in the maternal
the two Hageman
trait
when
from the 3.3-kb
in the human
In order
and
performed
which
two
subjects.
characterization
studies
in
shown),
of the
present
ofthe
to half
not
including
activity.
are
intensity
deficiency
indicate
reduced
and
in addition
to the Hageman
trait
and four females,
all of the paternal
FXII
The
FXII
arrows
was
RFLP
members
of both the paternal
and maternal
lineages.
The levels of FXII
activity
in polymorphic
and
morphic
subjects
of the paternal
lineage
together
lower
also
.
The
were
doublebands
of
are present
length
of the
double
digestions
has located
the
ofthe
FXII gene
5’ FXII
cDNA
(Fig
are
100
-2.1
0
0
0
0
0
0
0
S
><
S
S
50
-1.1
IU#{149}3 Ill’4
IV#{149}1
IV#{149}2lll3
IlI#{149}4
I
S
S
Taq I
Fig 2.
Taql RFLP detection.
DNA samples
from Hageman
subjects
(IV-i , IV-2)
and their
parents
(111-3. lIl-4)
have
digested
with Taql (A and B). Southern
blots were hybridized
3’ FXII cDNA (A) and to a 5’ FXII eDNA (B).
activity
assays.
Distribution
of FXII
activity
in polymorphic
(#{149})
and
nonpolymorphic
(0)
subjects
of the
paternal
lineage;
0.
unrelated
controls.
Fig
trait
been
to a
3.
FXII
clotting
S
S
S
8
o
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
FACTOR
GENE
XII
ALTERATION
IN HAGEMAN
Kb
TRAIT
i 423
Kb
Kb
1.7-
11-.
4-,
7III-3
Ill4
BamHI
present
and
to the
by
BgI II
within
450
intronic
pairs
at
additional
5’
FXII
indicating
long
base
sequences
TaqI
the
111-4
lll3
cDNA
that
The
sequences
bp
FXII
gene
bp)
exons
are
least
map
long,
with
in Fig
0.47-
5’
thus
lll-3 lll-4
relatively
of the
exonic
A TaqI RFLP
within
the gene
in two brothers
carrying
the
found in I 1 members
of their paternal
the identification
of the heterozygous
strating
that
the
autosomal
has been
trait and
lineage,
condition
pattern
of
inheritance
of
the two
are double
for
two
different
FXII
maternal
mutations
of gene
expression,
potential
and
different
lesions
as
within
of
TaqI
used
Taql
FXII gene
to the high
precise
This
of
polymorphism
demonstrated
the
gene
could
be
were
patterns
has
with
mutations
ognition
polymorphic
tetranucleotides
give
rise
site
to nonsense
in a gene
(TCGA)
additional
in
codons.
region
the
The
rich
in
coding
localization
intronic
FXII
a
splice
junction
mutation,
as
is
diseases.
Anderson
for revising
the manuscript.
if
a
as
several
a single
can be
TaqI
rec-
sequence
of the
sequences
Fijikawa
in the
blood
K, Kurachi
coagulation
K, Kisiel
cascade.
W: The
Adv
role
of serine
Enzymol
48:277,
1979
2.
Heimark
activation
3.
nature
4.
producing
Dr Barbara
EW,
proteases
Nature
excluded;
in the
REFERENCES
the
deletion,
possible
small
gene
alterations
is assumed,
a nonsense
mutation
cannot
of
only
If among
the
nucleotide
change
or
in genetic
been
available.
obtained
site
and from
than
portion
by a gene
intronic
found
We thank
enzymes.
in fact,
an
(lll-3)
ACKNOWLEDGMENT
of the
rather
obtained
polymorphic
TaqI
a polymorphic (111-4) subject
have
been
digested
with
BcIl/
Taql and Ncol/
Taql. The hybridization
has been performed
with the 5’ portion
of
eDNA.
(B) Schematic
representation
of restriction
sites
in a
portion
of the
FXII
gene.
B. BcIl; N. Ncol; P. PstI; T, Taql;
T,
polymorphic
site; 1 . 5’ eDNA;
2. 3’ eDNA.
Dotted
boxes indicate
the
tentative
position
of part of the exonic sequences.
1 . Davie
a
the
a nonpolymorphic
the
mutation
enzyme
in
of
from
in
with
lesion
localized
sequence
normal
Localization
suggests
lesions.”
is not produced
by the
restriction
that
gene
exons
of known
probe
this
than
by the 5’ cDNA
probe.
However,
due
of intronic
sequences
in the 5’ region,
a
assignment
genomic
that
infrequent
association
restriction
other
is
recognized
proportion
suggest
less
a
of two
polymorphism
its
a FXII
TaqI
polymorphism
with
presence
may
intragenic
be
to detect
individuals
5.
(A) DNAs
frequently
suggest
variant.
successfully
and
impairment
The
be
and
could
the
FXII.
family
TaqI
activity
site
a meaningless
The
this
paternal
complete
could
individuals
of FXII
additional
same
alteration
unrelated
reduction
the
of
Fig
gene.
heterozygous
Both
the
by
absence
gene
suspected.
The absence
lesions.
cause
indicated
material
asymptomatic
normal
gene
should
B
FXII
that
subjects
N
thus enabling
and demon-
only in the paternal
of parents
and indicates
trait
BN
identiis also
deficiency
is monogenic
recessive.
The presence
of the polymorphism
lineage
excludes
the consanguinity
Hageman
NcoI/TaqI
N
B
for FXII
Hageman
lll-3 lll-4
A BcII/TaqI
SB.
DISCUSSION
fled
-0.4
.a
4”
recognized
4 kb
of part
-0.7
site,
localized
region
interspersed
position
in the
polymorphic
long
is at
are
tentative
is indicated
3’ to the
I ,200
The
(350
short
introns.
(bp)
least
site.
-1.2
Fig 4.
Exclusion
of gene
deletion.
BamHl and Bglll DNA
digest
from
parents
(lll-3.
lll-4)
of the Hageman
trait subjects
have
been hybridized
to both
cDNA probes.
RL,
of
286:456,
K,
Fujikawa
coagulation,
K,
fibrinolysis
Davie
and
Ratnoff
OD:
Haemost
A familial
fraction
Veltkamp
zygotes
for
II,
factor
17:181,
trait
of plasma.
Hemker
VIII,
1965
HC,
IX
and
on
characterized
I Lab
Clin
Surface
formation.
Loeliger
EA:
XII
deficiency.
44:91
hereditary
of
5, 1954
Detection
of hetero-
Thromb
Diath
Hageman
inherited
trait
as an
(suppl)
6. Bennett B, RatnoffOD,
Holt IB, Roberts HR:
XII deficiency):
A probable
second genotype
dominant
the
by deficiency
Med
(factor
autosomal
EW:
kinin
1980
Margolius
A, Ratnoff
OD:
Observations
of Hageman
trait.
BlOOd I 1 :565, 1956
clot-promoting
5.
Kurachi
blood
characteristic.
Blood
40:412,
1972
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
i 424
BERNARDI
7.
Cool
DE,
Edgel
CIS,
Louie
GV,
Zoller
MacGillivray
RTA: Characterization
of human
factor XII cDNA. I Biol Chem 260:13666,
1985
8.
Tripodi
M, Citarella
F, Guida
5, Galeffi
Ml, Brayer
GD,
blood coagulation
P. Fantoni
R: cDNA
sequence
coding
for human
coagulation
(Hageman).
Nucleic Acids Res 13:3941,
1986
9. Panicucci
F: II Laboratorio
dell’Emostasi.
Pisa,
l985,p
127
A, Cortese
factor
Italy,
XII
Pacini,
10. Bernardi
Marchetti
G,
Gene deletion
22:305, 1985
1 1. Gitschier
Goralka
TM,
mutations
in
1985
ET AL
F, Del Senno L, Barbieri
F, Buzzoni D, Gambari
R,
Conconi
F, Panicucci
F, Positano
M, Pitruzzello
5:
in an Italian
haemophilia
B subject.
I Med Genet
I, Wood
WI, Tuddenham
EGD, Shuman
MA,
Chen EY, Lown RM: Detection
and sequence
of
the FVIII gene of haemophiliacs.
Nature
3 15:427,
From www.bloodjournal.org by guest on June 11, 2017. For personal use only.
1987 69: 1421-1424
Factor XII gene alteration in Hageman trait detected by TaqI restriction
enzyme
F Bernardi, G Marchetti, P Patracchini, L del Senno, M Tripodi, A Fantoni, S Bartolai, F Vannini, L Felloni
and L Rossi
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